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PRMT5-CATALYZED ARGININE METHYLATION OF NF-kappaB p65 IN THE ENDOTHELIAL CELL INDUCTION OF PRO-INFLAMMATORY CHEMOKINES

Harris, Daniel Pellerin
http://orcid.org/0000-0002-1852-0683
http://rave.ohiolink.edu/etdc/view?acc_num=case1449579234

Abstract Details

2016, Doctor of Philosophy, Case Western Reserve University, Physiology and Biophysics.
Inflammatory agonists differentially activate gene induction of chemokines in endothelial cells (EC). The molecular mechanisms that produce distinct chemokine induction profiles following EC activation are incompletely understood. The chemokines CXCL10/IP-10 and CXCL11/I-TAC both facilitate Th1-type leukocytes recruitment to inflammatory lesions. CXCL10 and CXCL11 are strongly induced by IFN-gamma (IFNg). However, CXCL11 expression is not triggered by TNF-a (TNF) exposure, whereas TNF potently induces the CXCL10 promoter. Both genes display strong synergistic expression upon simultaneous exposure to TNF and IFNg. In these studies we show that the arginine methyltransferase PRMT5 is critical in the expression of CXCL10 by TNF and CXCL11 in response to co-stimulation with TNF and IFNg, but that PRMT5 operates through distinctive mechanisms to achieve activation of these promoters. Chromatin immunoprecipitation experiments revealed that proteins containing the symmetrical dimethylarginine (SDMA) modification catalyzed by PRMT5 are found on each promoter following stimulation with pro-inflammatory agents. Using immunoblotting and mass spectrometry approaches we found that NF-kappaB p65 (p65), a key transcription factor critical for both CXCL10 and CXCL11 induction, contains PRMT5-catalyzed dimethylarginine. PRMT5-mediated methylation is required for p65 association with each promoter. A series of experiments revealed that methylation of different arginine residues on p65 is required for induction of CXCL10 versus CXCL11. CXCL10 induction requires methylation of Arg30 and Arg35 in response to TNF. In contrast, methylation of p65 at Arg174 is necessary for CXCL11 induction in EC costimulated with TNF and IFNg. Arg30, Arg35, and Arg174 are located in the rel homology domain of p65 but are present in regions associated with different biochemical functions. Arg30 and Arg35 are part of the DNA-binding domain. Methylation of these residues probably enhances p65 DNA binding affinity. Arg174 is found in a domain involved in protein-protein interactions. Therefore, methylation of Arg174 is likely to promote an association between p65 and a coactivating component of the CXCL11 transcription complex. In conclusion, I show that PRMT5 is a central player in chemokine gene expression by catalyzing the methylation of multiple arginine residues in p65. Methylation of p65 at specific sites comprises a code that enables unique chemokine gene expression profiles in EC.
Thomas Nosek, Ph.D. (Committee Chair)
Paul DiCorleto, Ph.D. (Advisor)
Cathleen Carlin, Ph.D. (Committee Member)
George Dubyak, Ph.D. (Committee Member)
Paul Fox, Ph.D. (Committee Member)
Mukesh Jain, M.D. (Committee Member)
167 p.
Biochemistry; Cellular Biology; Molecular Biology; Physiology
PRMT5; arginine methylation; SDMA; NF-kappaB; p76; post-translational modification; endothelial cell; inflammation; CXCL10; IP-10; CXCL11; I-TAC; TNF-a; IFNg; transcription

Recommended Citations

Citations

  • Harris, D. P. (2016). PRMT5-CATALYZED ARGININE METHYLATION OF NF-kappaB p65 IN THE ENDOTHELIAL CELL INDUCTION OF PRO-INFLAMMATORY CHEMOKINES [Doctoral dissertation, Case Western Reserve University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=case1449579234

    APA Style (7th edition)

  • Harris, Daniel. PRMT5-CATALYZED ARGININE METHYLATION OF NF-kappaB p65 IN THE ENDOTHELIAL CELL INDUCTION OF PRO-INFLAMMATORY CHEMOKINES. 2016. Case Western Reserve University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=case1449579234.

    MLA Style (8th edition)

  • Harris, Daniel. "PRMT5-CATALYZED ARGININE METHYLATION OF NF-kappaB p65 IN THE ENDOTHELIAL CELL INDUCTION OF PRO-INFLAMMATORY CHEMOKINES." Doctoral dissertation, Case Western Reserve University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=case1449579234

    Chicago Manual of Style (17th edition)

case1449579234
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This open access ETD is published by Case Western Reserve University School of Graduate Studies and OhioLINK.